1. Field of the Invention
The present invention relates to a monoclonal antibody for isolating and/or identifying mesenchymal stem cells.
2. Related Prior Art
In addition to stem cells for haematopoietic cells, stem cell-like cells, which constitute precursors of non-haematopoietic tissues, are also present in bone mar-row. These precursors of non-haematopoietic tissues were originally termed, inter alia, tissue culture dish-adherent (plastic-adherent) cells and are more recently being termed either mesenchymal stem cells or bone marrow stroma cells (MSCs).
In addition to their being of interest because of their multipotency in regard to differentiation, these cells are also of interest, for example, for their possible use in cell therapy and gene therapy.
The mesenchyme is the embryonic connective tissue, i.e. it is the multi-potent parental tissue for all forms of connective tissue and supporting tissue, for smooth musculature and for skeletal and cardiac musculature. Mesenchymal stem cells can be obtained and isolated from the bone marrow of adult humans. They are multipotent and contribute to the regeneration of bones, cartilage, ligaments, muscles, fat tissue and stroma.
The antibody W7C5 has been described by Giesert et al., “The mono-clonal antibody W7C5 defines a novel surface antigen on hematopoietic stem cells”, Annals of the New York Academy of Sciences 938: 175-183, as being a new marker for haematopoietic stem cells. At the same time, it was also shown in this publication that mesenchymal stem cells also express this marker. More recent studies show that this antibody recognizes the CD109 epitope on CD34+/CD38-stem cell populations.
Lagasse et al., “Purified hematopoietic stem cells can differentiate into hepatocytes in vivo”, Nat Med, 11: 1229-1234 (2000), Kopen et al., “Marrow stromal cells migrate throughout forebrain and cerebellum, and they differentiate into astrocytes after injection into neonatal mouse brains”, Proceedings of the National Academy of Science USA, 96: 10711-10716 (1999) and Brazelton et al., “From marrow to brain: expression of neuronal phenotypes in adult mice”, Science 290: 1775 1779 (2000), showed that mesenchymal stem cells which had been isolated from bone marrow were also able to differentiate into non-mesenchymal cells such as liver cells, neuronal cells and glial cells. In addition to this, it has only recently been shown that mesenchymal stem cells from adult human bone marrow are able to differentiate into neural cells in vitro. Woodbury et al., “Adult rat and human bone marrow stromal cells differentiate into neurons”, J. Neurosci. Res. 61: 364 370 (2000), showed that, in the presence of dimethyl sulphoxide (DMSO) and β-mercaptoethanol (BME), it was possible to differentiate mesenchymal stem cells into cells which expressed neurofilament and neurone-specific enolase. Other research groups reported the use of epidermal growth factor and brain-derived neurotrophic factor (BDNF) to differentiate stroma bone marrow cells into nerve cells which expressed nestin, glial fibrillary acidic protein (GFAP) and neurone-specific nuclear protein (Neu N).
The fact that mesenchymal stem cells are also able, under certain conditions, to differentiate into nerve cells means, inter alia, implicate the need to be able to distinguish these mesenchymal stem cells from neuronal precursor cells.
These neuronal precursor cells (=neural progenitor cells, in the following termed NPC) are to be found in the central nervous system. They also express nestin and are able to differentiate into neurones, astrocytes and oligodendrocytes.
Neuronal precursor cells are CD133-positive; this cell surface marker was originally found on haematopoietic stem cells. However, it has recently been shown that this marker is also expressed by nervous tissue and skeletal muscle tissue. For these reasons, this marker is not suitable, on its own, for the purpose of differentiating different stem cells or precursor cells.
Despite the great interest in mesenchymal stem cells, up to now no de-fined protocol for isolating and expanding the cells in culture does exist. Most experiments have been directed towards cultures of mesenchymal stem cells, which latter have been possible to isolate particularly as a result of their adhering firmly to tissue culture dishes.
Since, furthermore, mesenchymal stem cells and, for example, neuronal precursor cells are considered as being homogeneous populations, particularly in regard to their morphology and their phenotype, there is need, for these reasons as well, to be able to distinguish at least between both these stem cells and precursor cells.
U.S. Pat. No. 5,837,539 discloses antibodies which are specific for cell surface determinants on human mesenchymal stem cells. In order to generate these antibodies, mice were immunized for several days with a variety of human bone marrow-derived mesenchymal stem cells.
These antibodies were used, in particular, for differentiating mesenchymal stem cells from haematopoietic cells. In addition to this, it was found that these antibodies also crossreacted with other non-mesenchymal cells.